The long-term goal of this project is the characterization of neurotransmitter receptor-mediated information transduction and its regulation, across neuronal membranes. The primary, but not exclusive, model systems under investigation are those for dopamine (DA) receptors. In order to characterize DA and related receptors at the biochemical and molecular levels and study their regulation, there are two major interrelated lines of research which are ongoing: 1) investigation of the cell biology, function and regulation of the receptors at the protein level; and 2) the molecular cloning of the receptor genes and investigation of gene structure and regulation in normal and pathophysiological states. 1. Cell Biology and Regulation of DA Receptors: Characterization of the functional and regulatory properties of D1 and D2 DA receptors on various neuroblastoma and cDNA-transfected cell lines was continued. The D1A receptor was shown to undergo agonist-induced desensitization in CHO cells that is partially cAMP-mediated and involves both functional uncoupling and down-regulation of the receptors. Both short and long isoforms of the D2 receptor (D2S and D2L) were also shown to undergo desensitization in CHO cells in response to agonist treatment. The D2S receptor was also down-regulated by agonist treatment whereas the D2L receptor was paradoxically up-regulated. Both D2 receptor isoforms were expressed in NG108-15 neuroblastoma cells and shown to couple to K+ channels, albeit through different G proteins. The D3 receptor was also demonstrated to couple to K+ channels in the NG108-15 cells. 2. Molecular Cloning of DA and Other Receptors: The distribution of the D1A and D1B receptors were mapped in the kidney. Both D1 receptor subtypes were sequenced in the spontaneous hypertensive rat (SHR) which exhibits defective kidney D1 receptors. No differences in sequence were found in comparison to control rats. Work continued on the cloning of a third D1-like receptor which apparently is linked to the stimulation of phosphatidylinositol turnover and mobilization of calcium. Transgenic knock-out experiments for several of the DA receptor subtypes were initiated. Two completely novel serotonin receptors were cloned and expressed. These were designated the 5-HT6 and 5-HT7 serotonin receptor subtypes. Several other cDNA clones encoding putative orphan G protein-linked neurotransmitter receptors were identified.